Human respiratory syncytial virus is the main cause of lower respiratory tract infections among infants and young children (refs. 1 to 3—a list of references appears at the end of the disclosure and each of the references in the list is incorporated herein by reference thereto). Globally, 65 million infections occur every year resulting in 160,000 deaths (ref. 4). In the USA alone, 100,000 children may require hospitalization for pneumonia and bronchiolitis caused by RS virus in a single year (refs. 5, 6). Providing inpatient and ambulatory care for children with RS virus infections costs in excess of $340 million annually in the USA (ref. 7). Severe lower respiratory tract disease due to RS virus infection predominantly occurs in infants two to six months of age (ref. 8). Approximately 4,000 infants in the USA die each year from complications arising from severe respiratory tract disease caused by infection with RS virus and parainfluenza type 3 virus (PIV-3). The World Health Organization (WHO) and the National Institute of Allergy and Infectious Disease (NIAID) vaccine advisory committees have ranked RS virus second only to HIV for vaccine development.
RSV infection in adults was initially considered a significant problem only in certain high-risk populations, such as the institutionalized elderly. However, evidence has been accumulating that the infection occurs frequently in previously healthy adults (ref. 9).
RSV infections in the elderly usually represent reinfections in those who have had many prior episodes. These infections have been reported to cause altered airway resistance and exacerbration of chronic obstructive lung disease.
In adults over 60 years old, RSV usually causes mild nasal congestion and may also result in fever, anorexia, pneumonia, brochitis and deaths (ref. 10).
The structure and composition of RSV has been elucidated and is described in detail in the textbook “Fields Virology”, Fields, B. N. et al. Raven Press, N.Y. (1996), in particular, Chapter 44, pp 1313–1351 “Respiratory Syncytial Virus” by Collins, P., McIntosh, K., and Chanock, R. M. (ref. 11).
The two major protective antigens of RSV are the envelope fusion (F) and attachment (G) glycoproteins (ref 12). The F protein is synthesized as an about 68 kDa precursor molecule (F0) which is proteolytically cleaved into disulfide-linked F1 (about 48 kDa) and F2 (about 20 kDa) polypeptide fragments (ref. 13). The G protein (about 33 kDa) is heavily O-glycosylated giving rise to a glycoprotein of apparent molecular weight of about 90 kDa (ref. 14). Two broad subtypes of RS virus have been defined A and B (ref. 15). The major antigenic differences between these subtypes are found in the G glycoprotein while the F glycoprotein is more conserved (refs. 7, 16).
In addition to the antibody response generated by the F and G glycoproteins, human cytotoxic T cells produced by RSV infection have been shown to recognize the RSV F protein, matrix protein M, nucleoprotein N, small hydrophobic protein SH, and the nonstructural protein 1b (ref. 17).
A safe and effective RSV vaccine is not available and is urgently needed. Approaches to the development of RS virus vaccines have included inactivation of the virus with formalin (ref. 18), isolation of cold-adapted and/or temperature-sensitive mutant viruses (ref. 19) and purified F or G glycoproteins (refs. 20, 21, 22). Clinical trial results have shown that both live attenuated and formalin-inactivated vaccines failed to adequately protect vaccines against RS virus infection (refs. 23 to 25). Problems encountered with attenuated cold-adapted and/or temperature-sensitive RS virus mutants administered intranasally included clinical morbidity, genetic instability and overattenuation (refs. 26 to 28). A live RS virus vaccine administered subcutaneously also was not efficacious (ref. 29). Inactivated RS viral vaccines have typically been prepared using formaldehyde as the inactivating agent. Murphy et al. (ref. 30) have reported data on the immune response in infants and children immunized with formalin-inactivated RS virus. Infants (2 to 6 months of age) developed a high titre of antibodies to the F glycoprotein but had a poor response to the G protein. Older individuals (7 to 40 months of age) developed titres of F and G antibodies comparable to those in children who were infected with RS virus. However, both infants and children developed a lower level of neutralizing antibodies than did individuals of comparable age with natural RS virus infections. The unbalanced immune response, with high titres of antibodies to the main immunogenic RS virus proteins F (fusion) and G (attachment) proteins but a low neutralizing antibody titre, may be in part due to alterations of important epitopes in the F and G glycoproteins by the formalin treatment. Furthermore, some infants who received the formalin-inactivated RS virus vaccine developed a more serious lower respiratory tract disease following subsequent exposure to natural RS virus than did non-immunized individuals (refs. 24, 25). The formalin-inactivated RS virus vaccines, therefore, have been deemed unacceptable for human use.
Evidence of an aberrant immune response also was seen in cotton rats immunized with formalin-inactivated RS virus (ref. 31). Furthermore, evaluation of RS virus formalin-inactivated vaccine in cotton rats also showed that upon live virus challenge, immunized animals developed enhanced pulmonary histopathology (ref. 32).
The mechanism of disease potentiation caused by formalin-inactivated RS virus vaccine preparations remains to be defined but is a major obstacle in the development of an effective RS virus vaccine. The potentiation may be partly due to the action of formalin on the F and G glycoproteins. Additionally, a non-RS virus specific mechanism of disease potentiation has been suggested, in which an immunological response to contaminating cellular or serum components present in the vaccine preparation could contribute, in part, to the exacerbated disease (ref. 33). Indeed, mice and cotton rats vaccinated with a lysate of HEp-2 cells and challenged with RS virus grown on HEp-2 cells developed a heightened pulmonary inflammatory response.
Furthermore, RS virus glycoproteins purified by immunoaffinity chromatography using elution at acid pH were immunogenic and protective but also induced immunopotentiation in cotton rats (refs. 31, 34).
Influenza virus infection is one of the most common causes of respiratory tract diseases. Typically, the disease results in a high fever, usually 100° F. to 103° F. in adults, often higher in children, and respiratory symptoms, such as sore throat, running or stuffy nose, as well as headache, muscle aches and extreme fatigue. In a typical year, influenza is associated with about 20,000 deaths in the US, and many more hospitalizations (CDC).
Influenza viruses are divided into three types, designated A, B and C. Types A and B are responsible for epidemics that occur almost every winter. Influenza viruses continually change over time by mutation, which is termed antigenic drift.
Influenza A viruses are classified into sub-types on the basis of two surface antigens, hemagglutinin (H) and neuraminidase (N). Three subtypes of the hemagglutinin (H1, H2, H3) and two sub-types of neuraminidase (N1, N2) are recognized among influenza A viruses that have caused widespread human diseases. Immunity to these antigens reduces the likelihood of infections and lessens the severity of the disease if infection occurs.
As a result of antigenic drift, major epidemics of respiratory disease caused by new variants of influenza continue to occur. Thus, the antigenic characteristics of the circulating strains provide the basis for selecting the virus strains included in each year's vaccine.
Although there are many actual and potential benefits of vaccines that combine antigens to confer protection against multiple pathogens, these combinations may have a detrimental effect on the immunogenicity of the individual components.
As described above, RSV and influenza virus infections are prevalent in the adult population and particularly the elderly and it would be desirable to confer protection against such infection by the administration of a single vaccine composition. However, any potential detrimental effect of combining immunogens suitable for conferring protection against both RSV and influenza virus in a single formulation are unknown.